The overall objective of this research is to determine the mechanism by which inorganic arsenic (As) increases human disease risks. The specific goal is to determine the mechanism by which As acts as an endocrine disrupter, which our laboratory first reported and has now demonstrated occurs with several nuclear hormone receptors via a unique mechanism distinct from that of other known endocrine disrupters. We hypothesize that As-induced endocrine disruption is one of the principal means by which it is able to influence the wide array of disease risks including various cancers, diabetes, cardiovascular disease and developmental problems to which it has been linked in epidemiology studies, and that these effects are a result of As targeting one or more critical regulatory steps that are shared by these receptors, leading in turn to a variety of patho-physiological consequences. We will examine As effects on glucocorticoid receptor (GR)-mediated gene expression in H4IIE and EDR3 rat hepatoma cells as our principal model system, focusing on the differential effects of As on GR signaling at low and intermediate As doses. Our Specific Aims are to: 1) Determine the effects of very low dose As (0.01-1 u.M, 0.75-75 ppb) to enhance GR-mediated gene regulation. We hypothesize that these effects are a result of As targeting the early steps in receptor activation between binding of hormone and activation of transcription. 2) Determine the effects of intermediate dose As (1-3 |aM, 75-225 ppb) to suppress hormone receptor-mediated gene regulation. We hypothesize that these effects are distinct from the enhancement seen at lower doses, and involve the intermediate to later steps of receptor-mediated transcription. 3) Determine the effects of endocrine disruption by As on two hormone-regulated and As-affected genes, TAT and GREB1, that are likely to be involved in As-associated disease processes. We will use a combination of confocal microscopy, molecular biology, biochemistry, and proteomics approaches to investigate these questions. The long-term goals of this project are to provide mechanistic insights that can be used for more effective science-based risk assessments, for predicting the specific patho-physiological consequences of As exposure, for assessing gene-environment, agent-agent and other interactions, for assessing specifically sensitive sub-populations at elevated risk, and for developing effective interventions for these As-exposed populations.